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Analysis of Strep Assay Fluorescence and ODs in Clone 2 Observations

This report presents the findings from the Strep assay conducted on Clone 2, focusing on normalized fluorescence and optical density (OD) readings. Observations indicate that some constructs like AIDA-1 showed the highest fluorescence relative to others, despite a lower OD compared to the negative control. Notably, uninduced constructs exhibited higher background noise, complicating comparisons between clones. Additional findings reveal that compounds VtaA11, Hag, Hia, and others exhibited fluorescence surpassing that of the positive control but had lower ODs. Issues such as cell clumping and background noise were also documented.

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Analysis of Strep Assay Fluorescence and ODs in Clone 2 Observations

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  1. Mgfp5 Strep Assay Clone 2

  2. Unnormalized Fluorescence

  3. ODs of clone 2

  4. Observations • 8 = upaG_short (avg fluor 55) • 14 = AIDA-1 (avg fluor 55) • (maybe) 6 = Hag AtD (avg fluor 49) • AIDA-1 had a lower OD than the negative control. But relative to other constructs, it had the highest fluorescence reading • Uninduced constructs had high background because it also had higher ODs than the induced constructs

  5. Comparing Clone 1 to Clone 2 Clone1 Clone 2 Readings from both clones for these 3 constructs were consistently higher than other constructs.

  6. Strep Assay for INP

  7. INP ODs (after 3rd wash) = pellet loss and hits

  8. Unnormalized Fluorescence background PBS fluorescence subtracted

  9. Observations • When cells still in 96-well block, usually pellet, but some wells had clumped scattered cells at bottom • VtaA11, Hag, Hia, upaG_short, eCPX had fluorescences higher than positive control, but all of them had smaller ODs compared the control OD • Cell junk present in cpg_L2 – after removal, fluorescence dropped significantly

  10. KILR strep assay unnormalized fluorescence background PBS fluorescence subtracted

  11. EILD strep assay unnormalized fluorescence background PBS fluorescence subtracted

  12. Needle scFv unnormalized fluorescence

  13. Gliadin scFv unnormalized fluorescence

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